Introduction: Macrophages reside within the interstitial compartment of the testis and form junction complexes with Leydig cells in vivo. We have previously shown that testicular macrophages produce a factor that stimulates testosterone secretion by Leydig cells while peritoneal macrophages have no effect. The purpose of the studies of this application is to investigate the development and regulation of this paracrine phenomenon. Hypothesis: Monocytes from bone marrow migrate to the testis at an important time of development and a subpopulation of these cells are then influenced by local testicular factors in interact with Leydig cells. Specific Aims: 1. determine if the initial appearance of macrophages in the testis or the development of junctions between Leydig cells and macrophages occurs during an important time in testicular development, 2. determine if factors from the local testicular environment are involved in the differentiation or regulation of macrophages. Methods: We will determine the stage of development that macrophages first appear in the testis using light microscopic immunocytochemical methods. To determine when junctional complexes form between the macrophages and Leydig cells we will study the testes from animals at various stages of development or postnatal maturation using electron microscopic techniques. We will employ retrovirus mediated gene-transfer studies to determine if stem cells in the bone marrow produce testicular macrophages. To determine if local testicular factors are capable of regulating or inducing the testis- specific phenotype, we will treat macrophages from the testis, blood (monocytes), peritoneum, lung and ovary with factors form testicular extracts, interstitial fluid or culture medium of various testicular cell types and then assay for testis-specific functions. Significance: These studies will contribute to an understanding of the regulation and differentiation of the interaction between macrophages and Leydig cells. Since these multidisciplinary studies are broad-based and employ both in vivo and in vitro methods, important information concerning the physiological relevance of testicular macrophages will be gained.